The invention relates to a novel cysteine protease having homology to interleukin-1xcex2 converting enzyme (ICE). The protease represents a member of a family of cysteine proteases termed LICE (Like ICE) which may be involved in modulating apoptosis.
Programmed cell death, also called apoptosis, is essential for the maintenance of tissue size and cell number homeostasis of multi-cellular organisms, and also plays an important role in the development of diseases such as cancer, autoimmune disease, viral infection, and neurodegeneration. The importance of apoptosis in controlling cell proliferation as well as differentiation was not addressed until recently (for review, see Ellis et al. Ann. Rev. Cell Biol. 7, 663-698 (1991); Williams and Smith, Cell 74, 777-779 (1993); Vaux et al. Cell 76, 777-779 (1994); Hoffman and Liebermann, Oncogene 9, 1807-1812 (1994); White, Genes Dev. 10, 1-15 (1996); Abbas, Cell 84, 655-657 (1996). Molecular study of apoptosis was initiated in the nematode Caenorhabditis elegans whose development is strictly controlled by at least 14 genes regulating apoptosis (Ellis, supra; Hengartner and Horvitz, Curr. Opin. Genet. Dev. 4, 581-586 (1994a). Among these genes, ced-9 was found to suppress apoptosis and shared homology with the mammalian proto-oncogene Bcl-2 (Hengartner and Horvitz, Cell 76, 665-676 (1994b)). Other nematode genes, including ced-3 and ced-4, were required for programmed cell death to occur, and loss-of-function mutation in these two genes resulted in survival of cells that would normally undergo cell death (Ellis and Horvitz, Cell 44, 817-829 (1986); Yuan and Horvitz, Development 116, 309-320 (1992).
Structural analysis of the cloned ced-3 gene revealed the structural/functional homology between this gene product and a mammalian enzyme called interleukin-1xcex2 converting enzyme (ICE, Yuan et al. Cell, 75, 641-52 (1993). ICE cleaves the pro form of interleukin-1xcex2 (Black et al. FEBS Lett. 247, 386-390 (1989); Kostura et al., Proc. Natl. Acad. Sci. USA 86, 5227-5231 (1989)) and defined a new class of cysteine proteases (Cerretti et al., Science 256, 97-100 (1992); Thornberry et al., Nature 356, 768-774 (1992); Molineaux et al., Proc. Natl. Acad. Sci. USA 90, 1809-1813 (1993). However, it appeared that cleavage of IL-1xcex2 was not the only function of ICE. In cultured cells, over-expression of either CED-3 or ICE induced apoptosis (Miura et al., 1993). When ICE was introduced into dorsal root ganglia neurons by microinjection, it also triggered programmed cell death (Gagliardini et al. Science 263, 826-828, (1994). These data strongly suggested that both CED-3 and ICE contained a protease activity that was important for the control of programmed cell death, and that ICE or ICE-like cysteine protease(s) were involved in controlling mammalian apoptosis.
Although ICE shares strong homology with CED-3, its role in apoptosis is not well defined. Mice deficient in ICE develop normally and show little abnormality in tissue homeostasis (Li et al. Cell 80, 401-411 (1995); Kuida et al. Science 267, 2000-2003 (1995), suggesting that this subset of proteolytic enzymes might exhibit functional redundancy, or that ICE may not be involved in the major apoptotic mechanism.
A number of genes encoding putative cysteine 35 proteases that share sequence homology with CED-3 and ICE have been described recently. ICE-related genes include Nedd2 or Ich1 (Kumar et al., Genes Dev 3, 1613-1626 (1994); Wang et al., Cell 78, 739-750 (1994), Tx, Ich2 or ICErelII (Faucheu et al., EMBO J 14, 1914-1922 (1995); Kamens et al., J Biol Chem 270, 15250-15256 (1995), ICErelIII (Munday et al., J Biol Chem 270, 15870-15876 (1995), Mch2 (Fernandes-Alnemri et al., Cancer Res 55, 2737-2742 (1995), CPP32xcex2 (Fernandes-Alnemri et al., J Biol Chem 269, 30761-30764 (1994); Tewari et al., Cell 81, 801-809 (1995), and Mch3 or CMH-1 (Fernandes-Alnemri et al., Cancer Res 5, 6045-6052 (1995b); Lippke et al., J Biol Chem 271, 1825-1828 (1996). ICErelIII has also been isolated as Ty (see PCT Application No. 96/04387). The mouse ortholog of CPP32xcex2 has also been reported (Juan et al. Oncogene 13, 749-755 (1996)).
The role of ICE-like proteases in controlling apoptosis suggests that they will be useful for regulating events surrounding programmed cell death. As apoptosis is likely to be important in disorders characterized by excessive cell death (e.g., autoimmune diseases, viral infections and nerve cell degeneration) and in disorders involving increased cell proliferation (e.g., cancer), it is an object of the invention to identify genes and their encoded polypeptides that are involved in apoptosis. It is a further object of the invention to identify additional ICE-related cysteine proteases for the purpose of identifying new genes and proteins that control apoptosis.
A novel member of a family of ICE-like cysteine proteases has been identified. The family of proteases is termed LICE (Like ICE) and the family member described herein is termed LICE3. Nucleic acid sequences encoding LICE3 and nucleic acid sequences which hybridize to LICE3 sequences as shown in FIG. 1 and remain hybridized under stringent conditions are encompassed by the invention. Vectors and host cells for the expression of LICE3-encoding nucleic acid sequences are also provided for.
LICE3 polypeptides and derivatives thereof which exhibit at least 60% homology with the LICE3 amino acid sequence as shown in FIG. 1 are also included. Methods of production of LICE3 polypeptides and antibodies or fragments thereof which specifically bind LICE3 are also considered part of the invention.
The observed homlogy between LICE3 and other ICE-related genes which are known to encode polypeptides that are involved in apoptosis, suggests that LICE3 is also involved in controlling apoptosis or programmed cell death. Therefore, compounds which block LICE3 activity may reduce the rate and/or extent of apoptosis. Methods for identifying compounds which interact with LICE3 and block its activity are encompassed by the invention. Methods for the treatment of disorders characterized by increased cell proliferation or increased apoptosis are also included.